OBJECTIVES: The mineralocorticoid receptor (MR), a hormone-activated transcription factor, besides its role in controlling hydroelectrolytic homeostasis, exerts pro-adipogenic and anti-thermogenic effects, inhibiting mitochondrial-uncoupling protein UCP1 expression in brown adipocytes. The aim of this study was to gain insight into the molecular mechanisms by which MR participates in such metabolic regulation. METHODS: We evaluated in vivo MR effects on cold-induced UCP1 expression in MR-overexpressing mice. Expression profiles of several transcriptional coregulators were analyzed during differentiation of the brown adipocyte T37i cell line. Given that UCP1 expression is inversely controlled by catecholamines/retinoic acid and corticosteroids, we investigated the mechanisms of MR's inhibitory effect on UCP1 transcription in T37i cells. Chromatin immunoprecipitation (ChIP) experiments enabled us to explore MR interaction with UCP1 promoter regions. RESULTS: Cold-induced UCP1 expression was blunted in the brown fat of MR-overexpressing mice. Along with induction of increasing mRNA levels for specific adipocyte markers during T37i differentiation, MR coactivator transcript levels significantly increased in intermediate states of differentiation, whereas expression of MR corepressors transiently increased approximately 2-fold. Such a simultaneous transient peak in coregulator expression is consistent with physiologically relevant cooperation occurring during brown adipogenesis. ChIP demonstrated that, after retinoic acid stimulation and aldosterone exposure, MR and PPARγ concomitantly bind to specific UCP1 promoter motifs. CONCLUSION: Our studies demonstrate that MR exerts a pivotal metabolic role by controlling energy expenditure, and provide novel information on how MR participates in the regulation of brown adipocyte function.